It is well known that in different tissues, dihydropyridines bind at nanomolar concentrations top a receptor and are able to block voltage- operated Ca2+ channels. These channels have been referred to as L-type Ca2+ channels. Voltage-activated Ca2+ channels open, and close, or gate according to molecular transition rates that are regulated by transmembrane voltage. However, these channels are found in several types of non- excitable cells but the physiological role of voltage-gated Ca2+ entry in these cells is not well understood. In addition to the L-type Ca2+ channel, n, P, and T-type Ca2+ channels have also been described in the literature. However, prior to our reporting none have been described as being "steroid sensitive". The long term goal of this proposal is to determine the presence of L-type Ca2+ channel alpha, subunit transcripts and their dependence upon the steroid, testosterone in Harderian gland homogenates. Based upon published cDNA sequences, we have designed PCR primers to two highly conserved regions (S5-S6) of the fourth domain common to all known Ca2+ channel mRNAs. We will use RT-PCR to amplify these Ca2+ channel mRNAs. Ca2+ channel mRNA will be detected and quantitated by Southern analysis of PCR products and verified by Northern analysis. Using this approach, we will establish the genomic/nongenomic steroidal transcriptional control of the L-type Ca2+ channel. Our short term goal is to kinetically characterize the testosterone sensitive, high affinity dihydropyridine receptor with regard to saturation isotherms using two additional dihydropyridines (i.e., nimodipine and PN200-110), Ca2+ dependence of ligand binding, the effects of known Ca2+ channel blockers (e.g., verapamil and diltiazem), and the size of the alpha1, dihydropyridine binding subunit. The classical mechanism of steroid hormone action involves entry of hormone into a cell where the hormone binds to and induces a conformational change in its high affinity cognate intracellular receptor protein. These events are believed to be dependent exclusively on hormone binding. Recently, gonadal and adrenal steroids have been known to exert some of their electrophysiological and behavioral effects through non-genomic mechanisms. This constitutes an emerging but controversial concept in neuroendocrinology.